7-deoxylincomycin and analogs and isomers thereof and process for making the same



United States Patent ABSTRACT OF THE DISCLOSURE Ac NH I SR on are prepared by dehalogenation of a compound of the formula:

I SR OH BRIEF SUMMARY OF THE INVENTION This invention relates to novel compounds and to process for preparing them, and is particularly directed to 7-deoxylincomycin, and analogs and isomers thereof and to processes whereby they and like compounds are produced.

The novel compounds of the invention can be represented by the following structural formula:

OH I

wherein R is alkyl of not more than carbon atoms, advantageously not more than 8 carbon atoms, cycloalkyl of from 3 to not more than 8 carbon atoms, and aralkyl of not more than 12 carbon atoms, advantageously not more than 8 carbon atoms; and Ac is the acyl radical of a 4-substituted-L-2-pyrrolidinecarboxylic acid of the formula HRz (A) Ra aw t K 7 on c-on 1? ll H521 Ii wherein R and R are alkylidene of not more than 20 carbon atoms, including methylene, advantageously not more than 8 carbon atoms, cycloalkylidene of from 3 to 3,435,025 Patented Mar. 25, 1969 not more than 8 carbon atoms, and aralkylidene of not more than 12 carbon atoms, advantageously not more than 8 carbon atoms; and R is hydrogen or HR Examples of alkyl of not more than 20 carbon atoms (R, HR and HR are methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetr-adecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof. Examples of cycloalkyl are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2 methylcyclopentyl, 2,3 dimethylcyclobutyl, 2- methylcyclobutyl, and 3-cyclopentylpropyl. Examples of aralkyl are benzyl, phenethyl, 3-pheny1propyl, and 1- naphthylmethyl. Examples of alkylidene, cycloalkylidene, and aralkylidene groups (R and R include methylene, ethylidene, propylidene, butylidene, pentylident, hexylidene, heptylidene, octylidene, n-onylidene, decylidene, undecylidene, dodecylidene, tridecylidene, tetra decylidene, pentadecylidene, hexadecylidene, heptadecylidene, octadecylidene, nonadecylidene, eicosylidene, and the isomeric forms thereof, cyclopropylidene, cyclobutylidene, cyclopentylidene, cyclohexylidene, cycloheptylidene, cyclooctylidene, 2-cyclopropylethylidene, 3-cyclopentylpropy1idene, benzylidene, 2-phenylethylidene, 3-phenylpropylidene, and l-naphthylmethylene.

The novel compounds of the invention, Formula I, as well as other related compounds, can be prepared by replacing by halogen, the 7-hydroxy of a compound of the Formula II to form a 7-halo compound of Formula III and deshalogenating the latter to form a compound of Formula I.

The deshalogenation is effected advantageously by mixing the starting compound of Formula III with zinc dust and heating. In the process Ac and R can be any radical which is non-reactive with zinc dust, but for preparing compounds of the invention Ac and R are as given above. For example, when a compound of Formula IIIA (Ac in Formula III is that of the acid of Formula A) is used as the starting compound, a novel compound of Formula IA is obtained. When compound IA is hydrogenated with a catalyst effective to saturate an olefinic double bond, or the starting compound (Formula IIIA) is similarly hydrogenated and then deshalogenated, a compound of Formula IB is obtained as a mixture of cis and trans epimers according to the formulas which, if desired, can be separated by counter current distribution or chromatography.

When R (H R in Formulas B, IB, HE, and IIIB is hydrogen, it can be replaced by suitable alkylation or like procedure. Advantageously, this replacement is effected by reacting the compound according to Formula B, IB, IIB, or IIIB, wherein R is hydrogen with an oxo compound (an aldehyde or a ketone) and hydrogenating the resulting adduct with a catalyst effective to saturate an olefinic double bond. Either platinum or palladium can be used as the catalyst. Suitable oxo compounds have the formula R R CO where R R C is the same as R given above. Examples of suitable oxo compounds are formaldehyde, acetaldehyde, propionaldehyde, butyraldehyde, acetone, isobutyl methyl ketone, benzaldehyde, phenylacetaldehyde, hydrocinnamaldehyde, acetophenone, propiophenone, butyrophenone, 3 methyl 4 phenyl-Z- butanone, Z-methyl-S-phenyl 3 pentanone, 3-cyclopentanepropionaldehyde, cyclohexaneacetaldehyde, cycloheptanecarboxaldehyde, 2,2 dimethylcyclopropylacetaldehyde, 2,2-dimethylcyclopropyl methyl ketone, cyclopentyl methyl ketone, cyclobutyl methyl ketone, cyclobutanone, cyclohexanone, 4-methylcyclohexanone, and the like.

The starting compounds of Formula II are prepared by acylating a compound of the formula wherein Z is a protective hydrocarbyloxycarbonyl group which is removable by hydrogenolysis, trityl, i.e., triphenylmethyl, diphenyl(p methoxyphenyl)methyl, bis- (p-methoxyphenyl)phenylmethyl, benzyl, or p-nitrobenzyl, with a Wittig agent, e.g., an alkylidenetriphenylphosphorane [see e.g., Wittig et al., Ber., 87, 1348 (1954); Trippett, Quarterly Reviews, XVII, No. 4, p. 406 (1963)]. Examples of hydrocarbyloxycarbonyl groups (Z) are tertiary-butoxycarbonyl; benzyloxycarbonyl groups of the formula wherein X is hydrogen, nitro, methoxy, chloro, or bromo, for example, carbobenzoxy, p-nitrocarbobenzoxy, pbromo-, and pchlorocarbobenzoxy; and phenyloxycarbonyl groups of the formula wherein X is hydrogen, allyl, or alkyl of not more than 4 carbon atoms, such as phenyloxycarbonyl, p-tolyloxycarbonyl, p-ethylphenyloxycarbonyl, and p-allylphenyloxycarbonyl and the like.

In carrying out this process the 4-oxo-L-2-pyrrolidinecarboxylic acid (Formula C) is added to a freshly prepared Wittig reagent. The Wittig reagents herein used can be generally represented by the following formula:

wherein R is as given above. These Wittig reagents are prepared by reacting an alkyl, cycloalkyl, or aralkyltriphenylphosphonium halide with a base such as sodarnide, or sodium or potassium hydride, or the sodium or potassium metalate of dimethyl sulfoxide and the like. For example, the elimination of hydrogen halide from alkyltriphenylphosphonium halide, produces alkylidenetriphenylphosphorane. [The preparation of phosphoranes is discussed in detail by Trippett, Quart. Rev. XVII, No. 4, p. 406 (1963)]. The reaction is generally carried out in an organic solvent, such as benzene, toluene, ether, dimethylsulfoxide, tetrahydrofuran, or the like, at temperatures between 10 C. and the reflux temperature of the reaction mixture. The thus-obtained product, a 4- alkylidene-, 4-cycloalkylidene-, or 4-aralkylidene-l-protected-L-proline which has the following formula:

is recovered from the reaction mixture in a conventional manner, generally by extraction from aqueous solutions of the reaction mixture. The crude product can be purified by conventional means, such as recrystallization, chromatography, or formation and recrystallization of easily formed derivatives such as amine salts of the amino acid, e.g., the dicyclohexy'lamine salt, and the like, and liberating the amino acids from such compounds. By hydrogenating an acid of Formula D in the presence of a catalyst, e.g., platinum, which is effective to saturate a double bond, but which is ineffective to effect hydrogenolysis, a compound of the following formula:

is obtained. Platinum deposited on a carrier, e.g., carbon or an anion exchange resin like Dowex-l, a cross-linked polystyrene trimethylbenzylammonium resin in the hydroxide cycle is suitable. If desired, the starting compounds of Formula V can be acylated with acids of Formula C, D, or E to form compounds IIC, IID, and HE, respectively. Compound IIC can then be converted to compound IID by treatment with a Wittig reagent and compound IID hydrogenated to compound IIE by the procedures given above. The hydrogenation, both of the acid D and the acylate IID, gives a mixture of cis and trans epimers which, if desired, can be separated by counter current distribution or chromatography. The starting acids of Formula B in which R is hydrogen are obtained when an acid of Formula D or E is subjected to hydrogenolysis over a palladium catalyst, e.g., palladium on carbon. Likewise, compounds of Formula IID and 11B are converted to compounds of Formula IIB in which R is hydrogen by the same process. The starting acids of Formula B in which R, is hydrogen as well as compounds of Formula IIB in which R is hydrogen can be converted respectively to compounds of Formulas B and IIB in which R; is HR by the procedures given above.

The starting acids of Formula A are obtained by treating an acid of Formula D with hydrogen bromide in acetic acid to remove the Z group and then replacing the N-hydrogen with an HR group by the procedure given above. Compounds of Formula ID and IIE are converted to compounds of Formula IIA and IIB, respectively, [by the same process.

Some of the starting compounds of Formula II are obtained biosynthetically. Lincomycin, methyl 6,8-dideoxy-6-(trans-l-methyl 4 propyl L 2 pyrrolidinecarboxamido) 1 thio D erythro-a-D-galacto-octopyranoside, is obtained as an elalboration product of a lincomycin-producing actinomycete according to U.S. Patent 3,086,912. It has the following structural formula:

wherein R and R are methyl and R H is propyl. Lincomycin B, methyl 6,8-dideoxy-6-(transl-methyl-4-ethyl- L 2 pyrrolidinecarboxamido)-1-thio-D-erythro-a-D- galacto-octopyranoside (Formula VI wherein R and R are methyl and R H is ethyl) also is an elaboration product of the same microorganism when cultured according to the procedure given in U.S. Patent 3,086,912. Lincomycin C, ethyl 6,8-dideoxya6-(trans-1-methyl-4- propyl L 2 pyrrolidinecarboxamido)-l-thio-D-erythroa-D-galacto-octopyranoside (Formula VI wherein R is ethyl, R H is propyl, and R is methyl) is obtained when the process of U.S. Patent 3,086,912 is carried out in the presence of added ethionine. Lincomycin D, methyl 6,8- dideoxy 6 (trans 4 propyl-L-2-pyrrolidinecarboxarnido)-1-thio-D-erythro-a-D-galacto octopyranoside (Formula VI wherein R is methyl, R H is propyl, and R is hydrogen) is obtained when the fermentation of U.S. Patent 3,086,912 is carried out in the presence of added oc-MTL, methyl 6-arnino-6,8-dideoxy-D-erythro-l-thio-u- D-galacto-octopyranoside, a compound obtained by the hydrazinolysis of lincomycin. Methyl 6,8-dideoxy-6- (trans-4-ethyl L 2 pyrrolidinecar-boxamido)-1-thio-D- erythro-a-D-galacto-octopyranoside (Formula VI wherein R is methyl, -R H is ethyl and R is hydrogen) is also produced when a-MTL is added to the fermentation of U.S. Patent 3,086,912. Similarly, lincomycin K, ethyl 6, 8- dideoxy-6-(trans 4 propyl L 2 pyrrolidinecarboxamido)-1-thio-D-erythro oz D galacto octopyranoside (Formula VI wherein R is ethyl, R H is propyl, and R is hydrogen) is produced when the fermentation of U.S. Patent 3,086,912 is carried out in the presence of added a-ETL, ethyl 6 amino 6,8 dideoxy-l-thio D- erythro-a-D-galacto octopyranoside, a compound obtained by the hydrazinolysis of lincomycin C. Ethyl 6,8-dideoxy-6-(trans-4-ethyl L 2 pyrrolidinecarboxamido)- l-thio-D-erythro a D galacto-octopyranoside (Formula VI wherein R is ethyl, -R H is ethyl, and R is hydrogen) is also obtained when OL-ETL is added to the fermentation of U.S. Patent 3,086,912. The above-described N-desmethyl products which are obtained when oz-MTL and u-ETL are added to the fermentation process of U.S. Patent 3,086,912 are examples of compound IIB wherein R is hydrogen. By the procedure described above replacement of the N-hydrogens produces compounds of Formula IIB wherein R equals HR e.g., methyl 6,8- dideoxy 6 (trans 1 ethyl 4 propyl-L-Z-pyrrolidinecarboxamido) 1 thio D erythro-a-D-galacto-octopyranoside, ethyl 6,8 dideoxy 6 (trans-l-methyl-4- ethyl-L-Z-pyrro1idinecarboxamido) 1 thio-D-erythrd u-D-galacto octopyranoside, ethyl 6,8 dideoxy-6-(transl-ethyl 4 ethyl-L-2-pyrrolidinecarboxamido)-l-thio-D erythro a D galacto-octopyranoside, and methyl 6,8- dideoxy-6-(trans-1-ethyl-4-ethyl L 2 pyrrolidinecarboxamido)-1-thio-D-erythro a D galacto-octopyranoside.

As the biosynthetic-produced lincomycins, as well as the amino sugars derived therefrom, are either methyl or ethyl thioglycosides, it is sometimes desirable to convert them to higher or lower glycosides. It is sometimes desirable to convert any of the compounds of Formulas I, II, III or V to higher or lower glycosides, This can elfectively be accomplished 'by reacting the compound to be converted with a mercaptan of the formula R SH wherein R is an alkyl group of not more than 20 carbon atoms, but one different from R and cyclizing the resulting dimercaptal. For example, compounds of Formulas I, I I, and III on reaction with a mercaptan of Formula R SH produce dithioacetals of the formula XII the process advantageously is applied to any of the starting products of Formula II, i.e., I'IA, IIB, I'IC, IID, and HE. The resulting products can be subjected to hydrazinolysis to form compounds of the following formula:

HT) i| '(OH 0H XIII which can be N-acylated as described above with acids of Formulas A, B, C, D, and E to provide compounds according to Formula X-II wherein X is hydroxy. The process can also be applied to the starting compounds of Formula V. For example, oz-MTL on treatment with ethyl mercaptan followed by cyclization as described above is converted to oc-ETL.

An alternative process for making compounds of Formula XII or Formula XIII is to brominate the starting material (Formulas I, II, III, or V) and then react the product with a merca'ptan according to the following sequence:

The starting compound, XIV, is dissolved in water as a soluble salt, e.g., the hydrochloride, and bromine added with cooling advantageously to between about l and 20 C. It is sufiicient if the aqueous solution is cooled to about 05 C. and the bromine added dropwise. The stoichiometric amount of bromine is 1 mole for each mole of starting compound, though more or less can be used. Advantageously a slight excess, say from 5 to excess of bromine is used. The bromine initially replaces the RS group and the resulting intermediate hydrolyzes to the sugar in which the pyranose form XVa is in equilibrium with the aldose form XVb. In the presence of acid, e.g., hydrochloric acid or other strong non-oxidizing acid such as p-toluenesulfonic acid and sulfonic acid type anion exchange resins, the mercaptan R SI-I reacts with the sugar XV to form the thioglycoside XVI. Concomitantly some dithioacetal may be formed which after separation can be cyclized as described above to form more of the desired thioglycoside XVI.

The 7-hydroxy group of the starting compounds of Formula III can be replaced by chlorine by heating with thionyl chloride. In the process, Ac and R can be any radical which is non-reactive with thionyl chloride but for preparing compounds of the invention, Ac and R are as given above. All that is necessary, however, to effect the substitution of the 7-hydroxy by chlorine is to mix the starting compound of Formula II or V, advantageously in the form of an acid-addition salt, e.g., the hydrochloride, with thionyl chloride, advantageously in the presence of an inert solvent, with mild heating, advantageously at reflux temperature, until the desired substitution of the 7-hydroxy the reaction is carried out in an inert atmosphere, e.g., under nitrogen. Carbon tetrachloride can be used effectively as the solvent vehicle but other inert solvents such as chloroform, methylene chloride, ethylene chloride, ether, benzene, and the like can be used. A satisfactory procedure is to stir the reaction mixture at room temperature for a considerable period, say from about 1 to 18 hours or as long as necessary to obtain a reasonably clear solution and then to raise the temperature to between about and 100 C., for example, to the reflux temperature (77 C. for carbon tetrachloride). After the reaction is complete, usually after heating at reflux for about 1 to 5 hours, the reaction mixture is allowed to cool, advantageously under nitrogen. Any material that separates on the cooling is collected and dried. The solvent is removed by vacuum distillation at a pot temperature advantageously less than about 35 C. and the material which precipitates is collected and dried and treated with ethanol to convert any residual sulfite intermediates to the desired product. The collected material can then group by chlorine is effected. Advantageously,

be further purified by solvent extraction and/or recrystallization and can be recovered either as the free base or an acid addition salt.

The proportions of the reagents can be varied widely. Stoichiometrically, however, at least 3 moles of thionyl chloride is required for each mole of starting compound. Any larger amount can be used but ordinarily it is not necessary or desirable to use more than about a 10 fold excess. Advantageously, an excess of about 2 to 3 fold is used. The amount of solvent is not critical and can be varied widely in accordance with the practices in the art. Ordinarily from about 15 to about 30 volumes of solvent for each part of solid starting compound will suffice. The proportion of solvent to thionyl chloride, however, is important because of the solubility of the product in thionyl chloride. If the ratio of solvent to thionyl chloride (v./v.) is high, the desired product precipitates on cooling of the reaction mixture and the work up of the product is simplified. For example, with carbon tetrachloride a mixture of products precipitates directly on cooling the reaction mixture if the v./v. proportion of carbon tetrachloride to thionyl chloride is kept above about 10 to l.

The replacement of the 7-hydroxy group by halogen can be effected by mixing the starting compound of Formula II with Rydon reagent and heating. In the process Ac and R can be any radical which is non-reactive with Rydon reagent, but for preparing compounds of the invention Ac and R are as given above.

The mechanism by which Rydon reagent effects the substitution of the 7-hydroxy by halogen is not fully understood. It is believed, however, that the mechanism is such that a change in configuration results. Thus, a 7- hydroxy compound of the D-erythro configuration would yield a 7-halo compound of the L-threo configuration.

Rydon reagents are formed by the addition of halogen to triphenylphosphine or triphenylphosphite or addition of an alkyl halide to triphenylphosphite and can be represented by the formulas:

wherein X is halogen, e.g., chlorine, bromine, and iodine. Rydon et al., J. Chem. Soc. 2224 (1953); Ibid, 2281 (1954); Ibid 3043 (1956).

The Rydon reagent can be formed in situ by addition of halogen or methyl halide to a solution of the triphenylphosphine or triphenylphosphite in an inert solvent such as acetonitrile or dimethylformamide, or it can be isolated as a separate entity. In either case the reaction with the lincomycin or related compounds is effected by contacting the Rydon reagent therewith in an inert solvent, e.g., acetonitrile or dimethylformamide, until the desired substitution of the 7-hydroxy is obtained. The reaction takes place at ordinary temperature, though gentle heating can be effected if desired. Advantageously the temperature is maintained between about 20 C. and about 55 C. The product can be recovered from the reaction mixture by well known techniques such as filtration, solvent extraction, etc. The reaction mixture advantageously is heated with methanol to destroy any excess Rydon reagent, filtered to remove any solid such as triphenylphosphine oxide, formed in the reaction, and then treated to recover the product. The methanol can be added either before or after the filtration. Advantageously the treated and filtered reaction mixture is evaporated to dryness and purified by solvent extraction and/or chromatography.

The compounds of Formulas IA, IB, IIA, IIB, and V exist either in the protonated or non-protonated forms according to the pH of the environment. When the protonated form is intended the compound is qualified as an acid-addition salt and when the non-protonated form is intended it is qualified as the free base. The free bases can be converted to stable acid-addition salts by neutralizing the free base with the appropriate acid to below about pH 7.0, and advantageously to about pH 2 to pH 6. Suitable acids for this purpose include hydrochloric, sulfuric, phosphoric, thiocyanic, fluosilicic, hexafluoroarsenic, hexafluorophosphoric, acetic, succinic, citric, lactic, maleic, fumaric, pamoic, cholic, palmitic, mucic, camphoric, glutaric, glycolic, phthalic, tartaric, lauric, stearic, salicylic, 3-phenylsalicylic, S-phenylsalicylic, 3-methylglutaric, orthosulfobenzoic, cyclohexanesulfamic, cyclopentanepropionic, l,2-cyclohexane-dicarboxylic, 4-cyclohexanecarboxylic, octadecenylsuccinic, octenylsuccinic, methanesulfonic, benzenesulfonic, helianthic, Reineckes dimethyldithiocarbamic, cyclohexylsulfamic, hexadecylsulfamic, octadecylsulfamic, sorbic, monochloroacetic, undecylenic, 4-hydroxyazobenzene-4-sulfonic, octadecylsulfuric, picric, benzoic, cinnamic, and like acids.

The acid-addition salts can be used for the same purposes as the free base or they can be employed to upgrade the same. For example, the free base can be converted to a water-insoluble salt, such as the picrate, which can be subjected to purification procedures, for example, solvent extractions and washings, chromatography, fractional liquid-liquid extractions, and crystallization and then used to regenerate the free base form by treatment with alkali or to make a different salt by metathesis. Or the free base can be converted to a water-soluble salt, such as the hydrochloride or sulfate, and the aqueous solution of the salt extracted with various water-immiscible solvents before regenerating the free base form by treatment of the thus-extracted acid solution or converted to another salt by metathesis. The free bases of Formulas IA, IB, IIA, IIB, IIIA, IIIB, and V can be used as buffers or as antacids. The compounds of Formulas I, II, III and V react with isocyanates to form urethanes and can be used to modify polyurethane resins. The long chain compounds, i.e., where HR is alkyl of from 8 carbon atoms up, have surface active properties and can be used as wetting and emulsifying agents. The thiocyanic acid addition salt when condensed with formaldehyde forms resinous materials useful as pickling inhibitors according to US. Patents 2,425,320 and 2,606,155. The free bases also make good vehicles for toxic acids. For example, the fiuosilicic acid addition salts are useful as mothproofing agents according to US. Patents 1,915,334 and 2,075,- 359 and the hexafiuoroarsenic acid and hexafiuorophosphoric acid addition salts are useful as parasiticides according to US. Patents 3,122,536 and 3,122,552.

The close analogues of lincomycin, i.e., where R H is cis or trans alkyl of not more than 8 carbon atoms; R is methyl or ethyl; R is alkyl of not more than 8 carbon atoms, have antibacterial properties, and some are comparable or superior to lincomycin and can be used for the same purposes as lincomycin. The other analogues and isomers, including the components of Formulas IA and IB, have similar antibacterial properties but to a lesser degree and can be used for the same purposes as lincomycin where larger amounts are not objectionable.

The following examples are illustrative of the process and products of the present invention but are not to be construed as limiting. The parts and percentages are by weight and the solvent ratios are by volume unless otherwise specified.

EXAMPLE 1 7-de0xylincomycin A mixture of 10 g. of 7-chloro-7-deoxylincomycin, 300 ml. of water, and g. of zinc dust was stirred and heated at reflux for 15 hrs. The reaction mixture was cooled, filtered, and the filtrate freeze-dried to yield 8.0 g. of a white solid which was purified via chromatography over 1 kg. of silica gel as follows:

Fraction Ml. MeOH: CHCI Forerun 1, 200 1. 6 1-11 66 each Discard 12-24 66 each PREPARATION 1 7-chl0r0-7-de0xylinc0mwcin [methyl 7-chl0r0 6,7,8-tride0xy-6-(trans-1-methyl 4 propyl-L-2-pyrrolidinecarboxamido)-1-thi0-L-thre0-a-D galacto octopyranoside] SCH: on

(A) THE FREE BASE XIX A suspension of 221.0 g. (0.5 mole) of lincomycin hydrochloride in 5 l. of carbon tetrachloride was stirred well at 25 C. under nitrogen, 900 m1. of thionyl chloride was added all at once and stirring continued for 2 hr. During this period the solid dissolved and a clear solution was obtained. The reaction mixture was heated at reflux for 2 hr., the heat source then removed and nitrogen bubbled into the amber solution until the pot temperature dropped to 25 C. About 4 l. of liquid was removed via vacuum distillation at a pot temperature of less than 35 C. The yellow solid which precipitated during this distillation was collected and dried. This solid was dissolved in about 300 ml. of methanol, cooled at 25 C., made basic (pH 11) with dilute aqueous sodium hydroxide solution (2 N), diluted to about 1200 ml. with water and extracted well with ether. The ether extracts were combined, washed with a small amount of water, dried over anhydrous magnesium sulfate and filtered. On evaporation of an aliquot of the combined ether extract 7-chloro-7-deoxylincomycin free base is obtained as a yellow amorphous solid.

(B) THE HYDROCHLORIDE Addition of hydrogen chloride gas to the filtrate of Part A resulted in the precipitation of 7-chloro-7-deoxy- 11 lincomycin hydrochloride which was separated and recrystallized from ethanol and ethyl acetate. A 32% yield of white, crystalline 7-chloro7-de0xylincomycin hydrochloride solvated with approximately one molecular proportion of water was obtained.

Analysis.-Calcd. for C H ClN O S-HC1-H O: C, 45.18; H, 7.37; Cl, 14.82; N, 5.86; S, 6.70; H O, 3.77. Found: C, 44.70; H, 7.65; C], 14.27; N. 5.78; S, 6.45; H O, 3.85. [aJ +l30 (c.-0.9858 g./l() ml.). Activity: About 4 to 8 times lincomycin. Antibacterial spectrum: Same as lincomycin.

PREPARATION 2 7-chl0r0-7-de0xylincomycin free base The process of Preparation 1, Part A, was repeated except that methylene chloride was used in place of ether as the extracting solvent and the combined extracts were filtered and evaporated to dryness. There was obtained a 64% yield of 7-chloro-7-deoxylincomycin free base as a yellow amorphous solid. 15 grams of this amorphous solid was adsorbed on 30 grams of silica gel in methylene chloride, dried, and sifted onto a 3 inch diameter column of 1500 grams of silica gel. The column was then eluted with a mixture of methanol and chloroform in the proportions of 1 to 19, respectively, in 200 ml. portions after a 2 liter forerun. Fractions 26, 27, and 28 were pooled and evaporated to dryness yielding 1.04 grams of essentially pure 7-chloro-7-deoxylincomycin free base as an amorphous solid having an antibacterial spectrum of the same scope and magnitude as the 7chloro-7-deoxylincomycin hydrochloride of Preparation 1.

| t i T I t (ll-NUT- o no (A-l) CHLORINE SIBSTITUTIUN A suspension of 1 gram of lincomycin C hydrochloride in 25 ml. of carbon tetrachloride and 4.5 ml. of thionyl chloride was stirred under nitrogen for 2 hours at 25 C. A clear solution was obtained in about 15 minutes. The reaction mixture was then heated at reflux for 2 hours and evaporated to dryness under vacuum to give a yellow solid which was dried under vacuum at 40 C. for 18 hours. The product was then dissolved in about 15 ml. of warm ethanol, made basic with sodium hydroxide as in Preparation 1 and diluted to 300 ml. with water. The water solution was extracted 5 times with 100 ml. portions of ether. The ether extracts were pooled, dried over magnesium sulfate, filtered, saturated with hydrogen chloride gas, and evaporated to a brown residue which on being recrystallized two times from a mixture of ethanol and ethyl acetate (dissolved in a minimum amount of ethanol and ethyl acetate added to turbidity) yielded 200 mg. of 7-chloro-7-deoxylincomycin C hydrochloride as white crystals which had the same activity and spectrum as the 7-chloro-7-deoxylincomycin hydrochloride of Preparation 1.

(A fl) HALOGEN SI'BSTITUTION A solution of Rydon reagent was prepared by stirring a dry solution of 52.6 g. (0.2 M) of triphenylphosphine and 800 ml. of acetonitrile at 30 under nitrogen and 10 ml. (0.19 M) of bromine added over a 20-min. period.

After stirring for 10 min. more, 8.2 g. of lincomycin was added and the reaction stirred at 30 for 18 hr. A white solid was then present. The reaction was filtered and the solid discarded. Methanol ml.) was added to the filtrate and the solvents then evaporated under vacuum. The viscous residue was dissolved in 100 ml. methanol, diluted with 1800 ml. of water and extracted six times with 200 ml. portions of ether. The ether extracts were discarded, the aqueous phase made basic (pH 11) with aqueous KOH and then extracted four times with 200 ml. portions of methylene chloride. The extracts were dried and evaporated, leaving 11 g. of a yellow solid which was chromatographed over 1 kg. of silica gel, using methanolzchlorofor-m 1:9 (v./v.) as the solvent system. After a forerun of 1200 ml., 22 fractions of 56 ml. were collected. The last six (fractions 17-22) were pooled and evaporated to dryness yielding 2.8 g. of 7-bromo-7- deoxylincomycin. This was converted to the hydrobromide by dissolving in water, adding HBr to pH 1, filtering, and lyophilizing the filtrate. The hydrobromide had an Ot +114 (c. 0.9314, H 0) and the following analysis:

Calcd. for C H Br N O S: C, 39.28; H, 6.23; N, 5.09; S, 5.83; Br, 29.04. Found: C, 39.64; H, 6.19; N, 5.07; S, 6.04; Br, 28.59.

In place of bromine, there can be substituted other halogens. Chlorine, for example, yields 7-chloro-7-deoxylincomycin which is identical with the product obtained by chlorinating lincomycin with thionyl chloride. In place of triphenylphosphine there can be substituted triphenylphosphite. Also in that case a methyl halide can be used in place of the halogen. In place of the lincomycin, there can be substituted other lincomycins and analogs thereof. Thus, when lincomycin C is substituted for lincomycin, 7-bromo-7-deoxylincomycin C is obtained.

Lincomycin C is obtained by reacting lincomycin with ethanethiol (ethyl mercaptan) to form a diethyl dithioacetal and heating the reaction mixture in the presence of p-toluenesulfonic acid or heating to fusion. The following procedure is illustrative.

(11-1 0,55 DIDICUXYii-(TRANS-lMETHYL-4-PROPYLL-2- lYRROLII)IXECARBOXAMIDO) D ERYTHRO-D-GA- LA.CTO-ALDEIIYDO-OCTOSE DIETHYL DITHIOACETAL the temperature of the Well-stirred reaction mixture between 20 and 30 C. by cooling in acetone-Dry Ice. Solid potassium chloride was removed by filtration, and the solid washed well with chloroform. Additional chloroform was added to the filtrate (ca. cc.) and the mixture, stirred magnetically, was adjusted to pH 10 by the addition of aqueous sodium hydroxide (2 N). The chloroform layer was separated, the aqueous layer extracted thoroughly with chloroform, the combined extracts washed twice with water and dried over anhydrous sodium sulfate. Removal of the solvent at 30 C. in vacuo gave a semisolid residue, which on being crystallized from acetone, gave 5.41 gm. of 6,8-dideoxy-6-(trans-1-n1ethyl-4-propyl- L-Z-pyrrolidinecarboxamido) D-erythro-D-galacto-aldehydo-octose diethyl dithioacetal as colorless flattened needles, M.P. 130-132. Concentration of the mother-liquors gave additional material (1.50 gm.), M.P. 129131. (Total yield, 6.91 gm., 42.4%.)

Analysis.-Calcd. for c21H4 N2O5S21 C, H, 8.77; N, 5.81; S, 13.29%. Found: C, 52.38; H, 8.71; N, 5.93; S, 13.46%.

(13-2) CYCLIZATION TO LINCOMYCIN C (a) One part each of the diethyl dithioacetal of Part B1 and p-toluenesulfonic acid monohydrate were refluxed in 25 parts of acetonitrile until substantial antibacterial activity was obtained. The reaction mixture was cooled and evaporated to dryness and chromatographed on silica gel using a solvent mixture ethyl acetate, acetone and water in the ratio of 8:5:1, respectively. Fractions 102 through 131 showed antibacterial activity. Of these fractions, 105 through 125 were pooled, evaporated to dryness, and crystallized from acetone acidified with hydrochloric acid and recrystallized by dissolving in water and adding acetone to give crystals of lincomycin C hydrochloride, M.P. 149-153".

(b) The diethyl dithioacetal of Part B-l was heated to 260 for about 3 minutes and the odor of ethyl mercaptan was noted. The product on being chromatographed as in Part B2(a) yielded lincomycin C.

(C) PREPARATION OF LINCOMYCIN C BY FERMENTATION The lincomycin C hydrochloride, M.P. 151157 C. can also be prepared by the procedure described in Netherlands Patent 6,503,829 (Derwent 18,540) and A. D. Argoudelis et al., Biochemistry 4, 704-9 (1965).

(D) ALTERNATIVE METHOD FOR PREPARATION OF LINCOMYCIN C Lincomycin hydrochloride (8.85 g.-0.02 mole) was dissolved in 20 ml. of water, cooled at and stirred while adding bromine (3.52 g.0.022 mole) dropwise over a 1 minute period. Ethanethiol (25 ml.) was added and the mixture stirred at 25 for 2 hours. The clear, colorless, 2-phase system (ethanethiol is relatively insoluble in water) was cooled in an ice bath and hydrogen chloride gas bubbled in for about minutes. The lower, aqueous phase turned red. The reaction mixture was then extracted 3 times with 100 ml. portions of Skellysolve B and aqueous sodium hydroxide solution added to bring the aqueous phase to pH 11. The basic phase was extracted well with chloroform. The chloroform extracts were washed with saturated sodium chloride solution, dried, and evaporated under vacuum to yield 6.2 g. of a white solid. 4.8 g. of this solid was chromatographed over 800 g. of silica gel by the procedure of Preparation 2 using methanol-chloroform (1 :7, respectively) as the solvent system. After 800 ml. of forerun, 80 fractions of 25 ml. each were collected. Fractions 4058 were combined and evaporated to dryness and the residual solid recrystallized from acetone to yield 0.5 g. of material identical with the diethyl dithioacetal of Part B-l. Fractions 65-75 were combined, evaporated to dryness, and dissolved in a mixture of 5 ml. methanol and 400 ml. diethyl ether. Hydrogen chloride gas added and the white solid which precipitated was collected. On being recrystallized from aqueous acetone, 0.5 g. of lincomycin C hydrochloride, identical with that of Part C, was obtained.

(E) OTHER ALKYL 7 HALO-6,7,8 TRIDEOXY-6-(TRANS-1- METHYL 4 PROIYL-L 2-PYRROLIDINECARBOXAMI- DO) 1 TlIIO-L-THREO-a-D-GALACTO-OCTOPYRANO- SIDES By substituting the ethanethiol in Part B-1 and Part D of this preparation, with or without heating, by other alkyl mercaptans, for example, propyl, butyl, pentyl,

hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl mercaptans and the isomeric forms thereof; by cycloalkyl mercaptans, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2-methylyclopentyl, 2,3- dimethylcyclobutyl, 2 methylcyclobutyl and 3 cyclopentylpropyl mercaptans; or by aralkyl mercaptans, for

example, benzyl, phenethyl, 3-phenylpropyl, and 1 naphthylmethyl mercaptans, the corresponding alkyl, cycloalkyl, and aralkyl 6,8-dideoxy-6-(trans-1-methyl-4-propyl- L 2 pyrrolidinecarboxamido) 1 thio D erythroot-D-galacto-octopyranosides are obtained, which on treatment by the procedures of Preparation 1 and Part A are converted to the corresponding alkyl, cycloalkyl, and aralkyl 6,7,8-trideoxyand 7-halo-6,7,8-trideoxy-6-(trans- 1 methyl 4 propyl L 2 pyrrolidinecarboxamido)- l-thio L threo-a-D-galacto-octopyranosides. The compounds thus obtained (both the 6,8-dideoxy, the 6,7,8 trideoxy, and the 7-halo-6,7,8-trideoxy compounds) wherein alkyl is propyl, butyl, pentyl, and hexyl (obtained respectively when propyl, butyl, pentyl, and hexyl mercaptans are used) are elfective antibacterials, having the same spectrum as lincomycin.

PREPARATION 4 Methyl 7 chloro 6,7,8 trideoxy 6 (trams 1 alkyl- 4 butyl L 2 pyrrolidinecarboxamido) 1 thio- L-threo a D galacto octopymnosides A suspension of 116 mg. of methyl 6,8-dideoxy-6- (trans 1 ethyl 4 butyl L 2 pyrrolidinecarboxamido) 1 thio D erythro cc D galacto-octopyranoside free base in 3 ml. carbon tetrachloride and 0.7 ml. of thionyl chloride was stirred at 25 C. until a clear solution was obtained (about 15 minutes) and allowed to stand at 25 C. for 2 hours. The reaction mixture was then heated at reflux for 2 hours and then evaporated to dryness under vacuum to a yellow solid which was worked up as described in Example 3. The product, however, did not crystallize so the crystallizing solvents were evaporated to yield 17 mg. of methyl 7- chloro 6,7,8 trideoxy 6 (trans 1 ethyl 4 butyl- L 2 pyrrolidinecarboxamido) 1 thio L threoa-D-galacto-octopyranoside hydrochloride as a tan amorphous solid.

On substituting the cis epimer, there is obtained methyl- 7 chloro 6,7,8 trideoxy 6 (cis 1 ethyl 4 butyl- L 2 pyrrolidinecarboxamido) 1 thio L threo oz- D-galacto-octopyranoside hydrochloride having the same antibacterial spectrum.

On substituting the l-methyl analogs, methyl 7-chloro- 6,7,8 trideoxy 6 (cisand trans 1 methyl 4 butyl- L 2 pyrrolidinecarboxamido) 1 thio L threo a- D-galacto-octopyranosides hydrochlorides are obtained.

The cis and trans epimers used as starting materials in the above example were prepared as follows:

PART B.-4-BUTYLIDENE-1-CARBOBENZOXYJLI-PROLINE AND THE CYCLOHEXYLAMINE SALT THEREOF Sodium hydride (19 g.) as a 53% suspension in mineral oil was warmed with 350 ml. of dimethyl sulfoxide at a temperature of 75 C. until the reaction was complete (about minutes). After cooling to 32 C., 16.2 g. of butyl triphenylphosphonium bromide was added, and the resulting reaction mixture was stirred for 1 hour to insure complete reaction. A solution of 26 g. of 4- keto-l-carbobenzoxy-L-proline in 100 ml. of dimethyl sulfoxide was added, and the resulting mixture was heated at 70 C. for 3 hours. The reaction mixture was cooled to 25 C. and 1 liter of 2.5% aqueous potassium bicarbonate added. This mixture was washed twice with 700 ml. portions of ether and the ether was discarded after back extracting with 150 ml. of 2.5% aqueous potassium bicarbonate. The bicarbonate solutions were combined and acidified with 4 N hydrochloric acid. The acidified mixture was extracted with four 500-ml. portions of ether. The combined ether extracts were washed successively with 250 ml. of water, three 250-ml. portions of saturated aqueous sodium bisulfite, and 250 ml. of water, and dried over anhydrous sodium sulfate. Evaporation of the solvent under vacuum gave 24 g. of an oily residue which was 4-butylidene-l-carbobenzoxy- L-proline.

This residue was dissolved in 31 ml. of acetonitrile and treated with 18 ml. of dicyclohexylamine and refrigerated. The crystals were collected, washed with acetonitrile and dried in vacuo giving 21 g. (46.8%) of the crystalline dicyclohexylamine salt melting at 136- 140 C. After two recrystallizations from acetonitrile, an analytical sample was obtained which melted at 142l44 C. and had a rotation of [a] 4 (c.:O.99, CHCl Analysis.Calcd. for C l-l N O C, 71.86; H, 9.15; N, 5.78. Found: C, 71.69; H, 9.30; N, 5.74.

Ten grams of the dicyclohexylamine salt of 4-butylidene-l-carbobenzoxy-L-proline was shaken with ether and excess 5% aqueous potassium hydroxide until no solid remained. The layers were separated and each one was backwashed. The aqueous alkaline layer was combined with the backwash from the ether layer and acidified with 4 N hydrochloric acid. The mixture was repeatedly extracted with ether and the ether extracts were combined, dried over sodium sulfate, and evaporated in vacuo to give 6.3 g. (93%) of 4butylidene-l-carbobenzoxy-L- proline as an oil.

PART C.-4-BUTYL-1 CARBOBENZOXY IIPROLINE The oil from Part B was hydrogenated in 200 ml. of methanol over 2.1 g. of 10% platinum on Dowex-l catalyst under lbs. hydrogen pressure. The catalyst was removed by filtration and the filtrate evaporated to yield 6.3 g. of 4-butyl-l-carbobenzoxy-L-proline as an oil. The product contained about 2 parts cis-4-butyl11-carbobenzoxy-L-proline toeach part of trans-4-butyl-l-carbobenzoxy-L-proline.

If desired, the hydrogenation of the 4-ylidene group can be postponed to any later step, even to the final step, in the process.

By substituting the butyltriphenylphosphonium bromide of Part B by other substituted triphenylphosphonium bromides where the substituent is methyl, ethyl, propyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecylJoctadecyl, nonadecyl, and eicosyl and the isomeric forms thereof; cyclopropyl, cyclobuty], cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2-cyclopropylethyl, and 3- cyclopentylpropyl; benzyl, phenethyl, 3-phenylpropyl, and l-naphthylmethyl, the corresponding 4-alkylidene-, cycloalkylidene-, and. 4-aralkylidene-l-carbobenzoxy-L- prolines and the corresponding 4-alkyl-, 4-cycloalkyl-, and 4-ara1kyl-l-carbobenzoxy-L-prolines are obtained. For example, when the butyltriphenylphosphonium bromide is substituted by ethyl-, propyl-, isobutyl-, pentyl-, and hexyltriphenylphosphonium bromides there are obtained 4-ethylidene-1-carbobenzoxy-L-proline, 4-propylidene-l-carbobenzoxy-L-proline, 4-isobutylidene-1-carbobenzoxy-L-proline, 4-pentylidene-l-carbobenzoxy-L-proline, and 4-hexylidene-1-carbobenzoxy-L-proline, and cis 16 and trans 4-ethyl-l-carbobenzoxy-L-proline, 4-propyl-1- carbobenzoxy L proline, 4-isobutyl-l-carbobenzoxy-L- proline, 4-pentyl-l-carbobenzoxy-L-proline, and 4-hexy1- l-carbobenzoxy-L-proline.

PART DWMETIIYL 6-AMINO-6,s'DIDEOXY-1$21104)- EItYTlIIto-mD-GALACTO-OCTOPYRANOSIDE (a-MTL) A solution of 40 g. of lincomycin free base (US. Patent 3,086,912) in 20 ml. of hydrazine hydrate (98- 100%) was refluxed for 21 hours; excess hydrazine hydrate was then removed in vacuo under nitrogen at steam bath temperature, leaving a residue. The residue, a pasty mass of crystals, was cooled, acetonitrile was added, and the mixture was stirred until the crystals were suspended. The crystals were collected on a filter, washed with acetonitrile and with ether. The yield of White, crystalline a-MTL free base after drying in vacuo at room temperature was 21 g. (84% Recrystallization was accomplished by dissolving OL-MTL free base in hot dimethylformamide and adding an equal volume of ethylene glycol dimethyl ether.

Methyl 6 amino-6,8-dideoxy-l-thio-D-erythro-a-D-galacto-octopyranoside free base has a melting point of 225- 228 C., an optical rotation of [a] -l276 (c.=.768, water) and a pKa of 7.45.

Analysis.-Calcd. for C H NO S: C, 42.7; H, 7.56; N, 5.53; S, 12.66. Found: C, 42.6; H, 7.49; N, 5.75; 5, 12.38.

By substituting lincomycin by other alkyl or by cycloalkyl or aralkyl 6,8-dideoxy-6-(trans-l-methyl-4-propyl- L 2 pyrrolidinecarboxamido) 1 thio D erythroot-D-galaetooctopyranosides where alkyl is ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecy], tridecyl, tetradecyl, pentadecy'l, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof; cycloalkyl is cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2-methylcyclopentyl, 2,3-dimethylcyclobutyl, Z-methylcyclobutyl, and 3-cyclopentylpropy1; and aralkyl is benzyl, phenethyl, 3- phenylpropyl, and l-naphthylmethyl, the corresponding alkyl, cycloalkyl, and aralkyl 6-amino-6,8-dideoxy-1-thio- D-erythro-a-D-galacto-octopyranosides are obtained. For example, by substituting the lincomycin by ethyl-, propyl-, butyl-, pentyl-, and hexyl-6,8-dideoxy-6-(trans-l-methyl- 4 propyl L 2 pyrrolidinecarboxamido) 1 thio D- erythro-a-D-galacto-octopyranosides, ethyl 6-amino-6,8- dideoxy 1 thio D erythro a D galacto octopyranoside, propyl 6-amino-6,8-dideoxy-l-thio-D-erythrowD-galacto-octopyranoside, butyl 6-amino-6,8-dideoxy-lthio D erythro 0c D-galacto-octopyranoside, pentyl 6- amino 6,8 dideoxy l thio D erythro a D galacto-octopyranoside, and hexyl 6-amino-6,8-dideoxy-l-thio- D-erythro-a-D-galacto-octopyranoside are obtained.

If desired, the procedure of Part A can be applied to the compounds of Part D using the hydrochloride or other salt of a strong acid and the resulting 7-halo compounds processed by the ensuing steps of this example to the final products of the preparation. Also, if desired, the resulting 7-halo compounds can be deshalogenated by the procedure of Example 1 and the resulting 7-deoxy compounds processed by the ensuing steps of this preparation.

PART E.-.\[ETI1YL ds-DrDEoXY-d(l-CARBOBENZOXY- 4 BU'lYIrL-2 IYRROLIDINECARBOXAMIDO)-1-TIIIO- 1) ERYTIIRO-u.-D'GALACTO-OCTOPYRANOSIDE FREE BASE To a solution of 6.3 g. of 4-butyl-l-carbobenzoxy-L- proline (the oil from Part B) in m1. of distilled acetonitrile cooled to 0 there was added 3.46 ml. of triethylamine followed by 3.34 ml. of isobutyl chloroformate. The mixture was stirred at C. (:3") for 15 min. A solution of 6.2 g. of a-MTL free base from Part C in 85 ml. of water was added, and the reaction mixture was stirred at 0 C. for 0.5 and at 25 C. for 1 hr. The reaction product was then filtered and dried yielding 4.57 g. (37.7%) of methyl 6,8-dideoxy-6-(l-carbobenzoxy 4 butyl L 2 pyrrolidinecarboxamido) 1- thio-D-erythro-a-D-galacto-octopyranoside free base. The mother liquor was concentrated under vacuum and an additional 4.25 g. (35.2%) of product recovered. Recrystallization from acetonitrile produced crystals of methyl 6,8-dideoxy-6-( l-carbobenzoxy-4-butyl-L-2-pyrrolidinecarboxamido) 1 thio D erythro a D galactooctopyranoside free base melting at 194-196 C. A second recrystallization from acetonitrile afforded an analytical sample, M.P. 195.5200 C., [Odn-I-lll (c., 0.98, MeOH).

Analysis.-Calcd. for C N N O S: C, 57.75; H, 7.46; N, 5.13; S, 5.93. Found: C, 57.58; H, 7.16; N, 5.50; S, 6.07.

ROLIDINE'CARBOXAMIDO)-1-THIO D ERYTHRO-a-D- GALACTO-OCTOPYRANOSIDE HYDROCHLORIDE A solution of 7.8 g. of methyl 6,8-dideoxy-6-(1-carbobenzoxy 4butyl-L-2-pyrrolidinecarboxamido)-l-thio-D- erythro-ot-D-galacto-octopyranoside free base from Part E in 200 ml. of methanol was shaken over 2 g. of 10% palladium on carbon under 40 lbs. of hydrogen pressure for 17 hours. The catalyst was removed by filtration and the solution concentrated under vacuum. The residue was dissolved in a mixture of 20 ml. of acetone and 20 ml. of water and acidified with 6 N hydrochloric acid. Dilution with 4 volumes of acetone precipitated methyl 6,8-dideoxy 6 (4-butyl-L-2-pyrrolidine-carboxamido)-1-thio-D- erythro-a-D-galacto-octopyranoside hydrochloride which was collected by filtration and dried. The crystals, dried at 55 C. under vacuum, weighed 4.7 g. and melted at 188-l94 C. The analytical sample obtained by recrystallization from acetone melted at 197-199 C. and gave [a] +150 (water, c., 0.89).

Analysis.-Calcd. for C H N O S.HCI: C, 48.80; H, 7.96; N, 6.32; S, 7.24. Found: (corrected for 5.54% water) C, 48.58; H, 8.19; N, 6.04; S, 7.36.

This material possesses 8% of the antibacterial activity of lincomycin by S. [urea assay.

By substituting the oc-MTL by other alkyl or by cycloalkyl or aralkyl 6-amino-6,8-dideoxy-l-thio-D-erythro-a- D-galacto-octanopyranoside wherein alkyl is ethyl, propyl, butyl, pentyl, hexyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof; cycloalkyl is cyclopropyl, cyclobutyl, cycyclopentyl. cyclohexyl, cycloheptyl, cyclooctyle, 2-methylcyclopentyl, 2,3-dimethylcyc1obutyl, 2-methylcyclobutyl, and 3-cyclopentylpropyl; and aralkyl is benzyl, phenethyl, 3-phenylpropyl, and l-naphthylrnethyl, the corresponding alkyl, cycloalkyl, and aralkyl 6,8-diodeoxy-6-( l-carbobenzoxy 4-butyl-L-2-pyrrolidinecarboxamido)-l-thio-D-erythro-a-D-galacto-octopyranosides and alkyl, cycloalkyl, and aralkyl 6,8-dideoxy-6-(4-butyl-L-2-pyrrolidinecarboxamido l -thio-D-erythro-a-D-galacto-0ctopyranosides are obtained. For example, by substituting the a-MTL by ethyl, propyl, butyl, pentyl, and hexyl 6-amino-6,8-dideoxy 1-thio-D-erythro-a-D-galacto-octopyranoside, there are obtained ethyl 6,8-dideoxy-6-(1-carbobenzoxy-4- butyl L 2 pyrrolidinecarboxamido)-D-erythro-u-D- galacto-octopyranoside, propyl 6,8-dideoxy-6-(1-carbobenzoxy 4 butyI-L-Z-pyrrolidinecarboxamido)-1-thio- D erythro ot-D-galacto-octopyranoside, butyl 6,8-dideoxy 6 (l-carbobenzoxy-4-butyl-L2-pyrrolidinecarboxamido) 1 thio-D-erythro-a-D-galacto-octopyranoside, pentyl 6,8 dideoxy 6-(l-carbobenzoxy-4-butyl-L-2-pyrrolidineocarboxamido) 1 thio D-erythro-a-D-galactooctopyranoside, hexyl 6,8-dideoxy-6-(1-carbobenZoxy-4- butyl L-2-pyrrolidinecarboxamido)-1-thio-D-erythro-a- D galacto octopyranoside, ethyl 6,8 dideoXy-6-(4- butyl-L-2-pyrrolidinecarboxamido) 1 thio-D-erythroa-D-galacto-octopyranoside, propyl 6,8-dideoxy-6-(4- butyl L 2 pyrrolidinecarboxamido)-1-thio-D-erythroa-D-ga1acto-octopyranoside, butyl 6,8-dideoxy-6-(4-butyl- L 2 pyrrolidinecarboxamido)-1-thio-D-erythro-a-D- galacto octopyranoside, pentyl 6,8-dideoxy-6-(4-butyl-L-2- pyrrolidinecarboxamido) l-thio-D-erythro-a-D-galactooctopyranoside, hexyl 6,8 dideoxy-6-(4-butyl-L-2-pyrrolidinecarboxamido) 1-thio-D-erythro-a-D-galacto-octopyranoside.

By substituting the 4-butyl-1-carbobenzoxy-L-proline by other 4-alkyl-l-carbobenzoxy-L-prolines where the 4-alkyl is ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof; by 4-cycloalkyl-1-carbobenzoxy-L-prolines where 4-cycloa1kyl is cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl Z-methyl-cyclopentyl, 2,3-dimethylcyclobutyl, 4- methylcyclobutyl, and 3 cyclopentylpropyl; and by 4- aralkyl 1 carbobenzoxy-L-prolines where 4-aralkyl is benzyl, phenethyl, 3-phenylpropyl, and l-naphthylmethyl, the corresponding alkyl, cycloalkyl, and aralkyl 6,8-dideoxy-6-( 1-carbobenzoxy-4-alkyl, 4-cycloalkyl, and 4-aralkyl L Z-pyrrolidinecarboxamido)-l-thio-D-erythro-a- D-galacto octopyranosides, and the corresponding alkyl, cycloalkyl, and aralkyl 6 (4-alkyl, 4-cycloalkyl, 4- aralkyl L 2 pyrrolidinecarboxamido)-1-thio-D-erythro or D galacto-octopyranosides are obtained. For example, by substituting the 4-butyl-l-carbobenzoxy-L- proline by 4 methyl-, 4-ethyl-, 4-propyl-, 4-pentyl-, and 4-hexyl-1-carbobenzoxy-L-prolines, there are obtained methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8-dideoxy 6 (l carbobenzoxy-4-methyl-L-Z-pyrrolidinecarboxamido) 1 thio D erythro a D galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8 dideoxy-6-(1-carbobenzoxy-4-ethyl-L-2-pyrro1idinecarboxamido) 1-thio-D-erythro-a-D-galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8- dideoxy 6 (1-carbobenzoxy-4-propy1-L-2-pyrrolidinecarboxamido) 1 thio D-erythro-u-D-galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8- dideoxy 6-(1 carbobenzoxy-4-pentyl-L-2-pyrrolidinecarboxamido) 1 thio-D-erythro-a-D-galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8- dideoxy 6 (1 carbobenzoxy-4-hexyl-L-2-pyrrolidinecarboxamido) 1 thio D-erythro-a-D-galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8-dideoxy 6 (4-methyl-L-2-pyrrolidinecarboxamido) 1 thio D-erythro-a-D-galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8- dideoxy 6 (4-ethyI-L-2-pyrrolidinecarboxamido)-1- thio D erythro a-D-galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8-dideoxy-6-(4- propyl L 2 pyrrolidinecarboxamido)-1-thio-D-erythro a D galacto-octopyranosides; methyl, ethyl, propyl, butyl, pentyl, and hexyl 6,8-dideoxy-6-(4-pentyl-L-2- pyrrolidinecarboxamido) 1 thio-D-erythro-u-Dgalactooctopyranosides; and methyl, ethyl, propyl, butyl, pentyl,

and hexyl 6,8 dideoxy 6-(4-hexyl-L-2-pyrrolidinecarboxamido) 1 thio-D-erythro-a-D-galacto-octopyranosides.

If desired, the I-carbobenzoxy compounds prepared according to Part E can be halogenated by the procedures of Part A, followed by dehalogenation by the procedure of Example 1 if desired, and the resulting 7-halo and 7- deoxy compounds processed by the ensuing steps of the preparation to remove the l-carbobenzoxy group and to 19 substitute the proline nitrogen to give 7-halo or 7-deoxy products.

PART G-1..\1ETHYL fi.S-DIDEOXY-G-(l MEllTYL-l-lil 'lYL L 2 IYRROLIDINECARUOXAMIDO)-1-THIO-1)- A mixture of 2.0 g. of methyl 6,8-deoxy-6-(4-butyl-L-2- ERYTiIRO-CL-I)-GALACTO OCTOPYRANOSI DE HYDE 5 pyfrolidinecarboxamido) 1 thi D e -ythro D galacto.

(momma r r XXVII on. XXVIII N\ z J not A not COMTL COMTL R C4119 A solution of 2.0 g. of methyl 6,8-dideoxy-6-(4-butyl- L 2 pyrrolidinecarboxamido) l thio D-erythro-a-D- galacto-octopyranoside hydrochloride from Part F and 2.0 ml. of 37% formalin in 150 ml. of methanol was shaken over 500 mg. of palladium on carbon under lbs. of hydrogen pressure for 3.5 hrs. Removal of the catalyst by filtration and the solvent by distillation in vacuo yielded partially crystalline methyl 6.8-dideoxy-6- (l-methyl-4-butyl-L-Z-pyrrolidinecarboxamido) -1-thio-D- erythro-a-D-galacto-octopyranoside hydrochloride which by TLC (thin layer chromatography) on silica gel using a mixture of ethyl acetate, acetone, water (8:4:1) for elution and KMnO, solution for detection consisted chiefly of two materials, the cis and trans epimers of methyl 6,8-dideoxy-6-( l-methyl-4-butyl-L 2 pyrrolidinecarboxamido)-1-thio-D-erythro-a-D-galacto-octopyranoside hydrochloride in a ratio of about 3 to 2.

G-2.-SI-I1ARATIO.\ OF THE CIS AND TRANS FORMS BY CllltO.\1.-\TOGR.\111Y The methyl 6,8-dideoxy-6-(1-methyl-4-butyl-L-2-pyrrolidinecatboxamido) 1-thio-D-erythro-a-D-galacto-octopyranoside hydrochloride from Part G-l were dissolved in a mixture of methanol and methylene chloride (1:1) and 1.5 ml. of triethylamine added. To this solution was added 7 g. of silica gel and the solvent evaporated under vacuum leaving the antibiotic deposited on the silica gel which was sifted on top of a chromatographic column of 200 grams of silica gel packed with a solvent mixture consisting of ethyl acetate, acetone, water in a ratio of 8:4:1. The column was developed by eluting with the same solvent and 20 ml. portions were collected. Thin layer chromatography of each fraction showed that fractions 31-38, 310 mg, were essentially pure trans epimer, that fractions 49-74, 32 mg., were essentially pure cis epimer, and that fractions 39-48 consisted of a mixture of epimers. The latter could be further separated by repeated chromatography. Each epimer was dissolved in a few drops of dilute hydrochloric acid and the hydrochloride precipitated by addition of acetone. in this manner, there was obtained mg. of methyl 6,8-dideoxy-6- (trans-l-methyl-4-butyl-L-Z-pyrrolidinecarboxamido) 1 thio-D-erythro-a-D-galacto-octopyranoside hydrochloride,

M.P. -137" and about mg. of methyl 6,8-dideoxy- 6-(cis-l-methyl-4-butyl-L-2-pyrrolidinecarboxa mido) l thio-D erythro-a-galacto-octopyranoside hydrochloride, softening at 105 C. with further melting at -185 C.

The trans epimer recrystallized from the same solvent melted at 13914l C. and had the following analysis:

Analysis.-Calcd. for C19H35N20sS-HC1I C, H, 8.16; N, 6.13; S, 7.02. Found: (Corrected for 4.07% H O);C, 48.81; H, 8.54; N, 6.49; S, 6.67.

Similarly recrystallization of the cis epimer gave a product softening at 108 C. and further at about 189 C. (solvated) with the following analysis:

Analysis.Found: (Corrected for 4.95% water); C, 50.27; H, 9.00; N, 6.05; S, 6.65.

The trans epimer was about 2.2 times as active as lincomycin by S. lutea assay, about 2 times as active by the broth dilution assay, and 2.5 times as active in mice infected with S. aureus.

The cis epimer was about /2 to /3 as active as the trans epimer, being about equal to lincomycin.

octopyranoside hydrochloride from Part F, 1.5 ml. of acetaldehyde, 150 mg. of 10% palladium on carbon in 150 ml. of methanol was shaken under 32 lbs. of hydrogen pressure for 5.5 hrs. The catalyst was removed by filtration to give a residue consisting chiefly of the cis and trans epimers of methyl 6,8-dideoxy-6-(1-ethyl-4-butyl-L- 'l-pyrrolidinecarboxamido)-l-thio-D-erythro-a-D-galactooctopyranosicle hydrochloride.

1I-2.SEPARATION OF EPIMERS As described in Part G-2, the mixture of epimers of Part H-1 (2 g.) was chromatographed over 200 g. of silica gel using for elution a solvent system of ethyl acetate, acetone, water (824:1). Fractions 33-42 by TLC were pure trans-epimer and were combined, fractions 49-64 were essentially pure cis-epimer and were also combined. Fractions 43-48 were a mixture of the epimers which could be purified by rechromatography. Each epimer was dissolved in a few drops of dilute hydrochloric acid and the crystalline hydrochloride precipitated on dilution with a large volume of ether.

The crude trans epimer fraction of 415 mg. gave 340 mg. (15.4%) of crystalline methyl 6,8-dideoxy-6-(trans- 1-ethyl-4-butyl-L-2-pyrrolidinecarboxamido)-1 thio D erythro-ot-D-galacto-octopyranoside hydrochloride, M.P. 144-15l C. Recrystallization from dilute acetone raised the M.P. to 148-151 C.

The cis epimer fraction of 645 mg. afforded 300 mg. (14.1%) of crystalline methyl 6,8-dideoxy-6-(cis-l-ethyl- 4-butyl-L-2-pyrrolidinecarboxamido)-l-thio-D-erythro a- D-galacto-octopyranoside hydrochloride, M.P. 135-139 C. Recrystallization from dilute acetone gave crystals, M.P.134138 C.

The trans epimer isomer showed about 1-1.2 times the activity of lincomycin by the S. lutea assay, 2-4 times the activity of lincomycin against gram positive organisms, and 8 times or more the activity of lincomycin against gram-negative organisms. In mice against S. aureus the trans epimer was about 2 times as active as lincomycin. The cis epimer was about /2 as active as the trans epimer.

Separation of the cis and trans isomers is not a necessary step as the 7-chlor0 derivatives of the mixed epimers are useful per se. It is desirable, however, to keep the content of trans isomer high as this is the most active form. By carrying out the process with this in mind mixed epimeric products containing a ratio of trans and cis epimers of 3:1 to 1:5 can readily be obtainer. By substituting the formaldehyde and acetaldehyde of Parts G and H by other oxo compounds of the formula R R CO, for example, propionaldehyde, acetone, butyraldehyde, isobutyl methyl ketone, benzaldehyde, phenylacetaldehyde, hydrocinnamaldehyde, acetophenone, propiophenone, butyrophenone, 3-methyl-4-phenyl-2 butanone, 2 methyl-5- phenyl-3-pentanone, 3-cyclopentanepropionalaldehyde, cyclohexaneacetaldehyde, cycloheptanecarboxaldehyde, 2,2- dimethylcyclopropaneacetaldehyde, 2,2 dimethylcyclopropyl methyl ketone, cyclopentyl methyl ketone, cyclobutyl methyl ketone, cyclobutanone, cyclohexanone, and 4-methylcyclohexanone, and using the appropriate alkyl,

' cycloalkyl, or aralkyl 6,8-dide0xy-6-(4-alkyl, 4 cycloalkyl-, or 4-aralkyl-L-2-pyrrolidinecarboxamido)-1-thio- D-erythro-tx-D-galacto-octopyranoside, there are obtained the corresponding alkyl, cycloalkyl, and aralkyl 6,8-dideoxy-6-(1-R R CH-4-alkyl, 4-cycloalkyl-, and 4-aralkyl-L- 2-pyrrolidinecarboxamido)-1-thio-D-erythro a D-galacto-octopyranosides which on treatment with thionyl chloride by the procedure of Part A gives the corresponding alkyl, cycloalkyl, and aralkyl, 7-chloro-6,7,8-trideoxy- 6-(1-R R CH-4-alkyl, 4-cycloalkyl-, and 4-aralkyl L-2- pyrrolidinecarboxamido)-l-thio-L threo a-D-galactooctopyranosides where R R CH- is propyl, isopropyl, butyl, and 4-methyl-2-pentyl; benzyl, phenethyl, 3-phenyl propyl, l-phenylethyl, l-phenylpropyl, l-phenylbutyl, 3- methyl-4-phenyl-2-butyl, and Z-methyl-S-phenyl-3-pentyl; 3 cyclopentylpropyl, 2 cyclohexylethyl, cycloheptylmethyl, 2 (2,2 dimethylcyclopropyl)-ethyl, l-(2,2-dimethylcyclopropyl)ethyl, 1 cyclopentylethyl, 1 cyclobutylethyl, cyclobutyl, cyclohexyl, and 4-methylcycl0- hexyl. By using formaldehyde and acetaldehyde or other alkanals, for example, propionaldehyde, butyraldehyde, valeraldehyde, or caproaldehyde with an alkyl 6,8-dideoxy-6-(4-alkyl-L 2 pyrrolidinecarboxamido)-1-thio-D- erythro-a-D galacto-octopyranoside where alkyl and 4- alkyl are methyl, ethyl, propyl, butyl, pentyl, or hexyl, preferred starting compounds of the formula 0H XXIX wherein X is hydroxy; R, HR and R are alkyl of not more than six carbon atoms, advantageously of not more than twelve carbon atoms in the aggregate; and the configuration is D-erythro, are obtained which on treatment with thionyl chloride or Rydon reagent by the procedure of Part A give compounds according to Formula XXIX where X is halogen; and which on further treatment with zinc dust by the procedure of Example 1 gives compounds of Formula XXIX wherein X is hydrogen; R, HR and R in both cases are alkyl as given above; and the configuration is believed to be L-threo. The 6,8-dideoxy, 6,7,8-trideoxy, and 7-halo-6,7,8-trideoxy compounds of Formula XXIX as characterized above are antibacterials and have the same spectrum of antibacterial activity as lincomycin. Representative compounds of Formula XXIX wherein X is hydrogen but which are believed to have the L-threo configuration are given in the following table:

TABLE I. COMPOUNDS OF FORMULA XXIX WHERE X IS HYDROGEN R HRl a Trans-ethyl Methyl. 4B do Cis ethyl Do. Trans-propyl D0. myem). 4D (7-deoxyall0linc0- .....do Cis-propyl Do.

rnyciu 4E (7-deoxylinco- ...do Trans-propyl Ethyl.

mycin E). 4F (7-deoxyallolincod0 Cispropyl Do.

myein E). G Trans-propyl. Methyl.

Cis-propyl Do. Trans-butyl Do. Cis-butyl. Do. Trans-propyl Ethyl. Cis-propyl Do. Trans-propyl. D0. Cis-propyl." D0. Trans-butyl D0. Cis-butyl- Do.

Trans-pentyl Methyl Cls-pentyl Do. Tians-butyl Ethyl. Crs-butyl. Do. Trans-pentyl Do. Cis-pentyl Do.

Cls-propyl D0. Trans-pentyl Ethyl. Cis-pentyl. Do. Trans-butyl Do. Cis-butyl D0. Tlans-penty Methyl Cis-pentyl. Do. 'Irans-propyl D0. Cis-propyl Do.

Trans-pentyL. Ethyl. Cispentyl. Do. Tians-pentyl. Do. Cispentyl Do.

Intermediates for the preparation of the above compounds correspond to the above table where (1) X is halogen; (2) R is hydrogen; (3) X is halogen and R is hydrogen; (4) X is halogen and R is carbobenzoxy; (5) X is halogen, R is carbobenzoxy, and HR and H in the 4-position are replaced by the ylidene group, R (6) X is halogen, R is hydrogen and HR and H in the 4-position are replaced by the ylidene group, R and (7) X is halogen, and HR and H in the 4-position are replaced by the ylidene group, R and it being understood that the configuration is believed to be L-threo. The corresponding compounds Where X is hydroxy having the D-erythro configuration are used as starting compounds.

When the lincomycin of Part A-2 of Preparation 3 is replaced by alkyl, cycloalkyl, or aralkyl 6-amino-6,8-dideoxy-l-thio D-erythro a-D-galacto-octopyranoside free base there are obtained compounds of the formula:

REE.

OH XXX SCH 0H XXXI A solution of 9.8 g. of lincomycin in 150 ml. of acetone is added to a solution of 9.8 g. of p-toluenesulfonic acid monohydrate in ml. of acetone with good stirring and avoidance of exposure of moisture. The mixture is stirred at ambient temperature for 1 hour, after which 100 ml. of anhydrous ether is added and stirring is continued in an ice-bath for 0.5 hour. The mixture is filtered and the solid is dried in vacuo at 50 0.; yield 13.35 g. (85.5%) of 3,4-O-isopropylidenelincomycin p-toluenesulfonate. An additional 1.15 g. (7.4%) can be recovered from the mother liquors by adding 350 ml. of anhydrous ether to the mother liquor from the previous filtering operation and chilling the solution for 1 hour. The 14.5 g. so obtained are suspended in 200 ml. of ether and shaken 23 vigorously with 125 ml. of potassium bicarbonate solution. The aqueous layer is back-extracted with two 100ml. portions of ether. The ether extracts are washed with 50 ml. of saturated sodium chloride solution and then filtered through anhydrous sodium sulfate. The ether is evaporated under vacuum, leaving 7.9 g. (73.1%) of 3,4-O-isopropylidenelincomycin which is dissolved in 25 ml. of ether acetate and concentrated to about to ml. The concentrate is allowed to stand at room temperature for several hours and then refrigerated overnight. The crystals are filtered from the solution and washed sparingly with cold ethyl acetate; yield 4.55 g. (42.2%) of 3,4-O-isopropylidenelincomycin having a melting point of 126-128 C., and an optical rotation of [04 101- 102 (c, 1, methylene chloride).

(3) TDEIIXDRO3,4-O-ISOlROPYLIDENELINCOMYCIN To a solution of 6 g. (0.0135 mole) of isopropylidenelincomycin in 75 ml. of pyridine was added 12 g. (excess) chromic oxide. The solution warms up about C. After one hour the mixture was added to a solution containing 250 ml. each of ethyl ether and ethyl acetate. This was then filtered and evaporated to a syrup, 8.4 g. This syrup was distributed in a SOD-transfer counter current distribution using the system, waterrethyl acetate:ethanol:cyclohexane (1:1:1:1). 7-dehydro-3,4-O-isopropylidenelincomycin was isolated as the peak fraction from tubes 330- 380, K:2.45.

Analysis.-Calcd. for C H N O S: C, 56.72; H, 8.16; N, 6.30; S, 7.21. Found: C, 56.37; H, 7.62; N, 6.51; S, 6.84.

(C) 3,4-O-ISOPROPYLIDENl-I-El'ILINCOMYCIN To 1.6 g. of Craig-pure 7-dehydro-3,4-O-isopropylidenelincomycin in 75 ml. of methanol was added 400 mg. of sodium borohydride. After 1.5 hr. this solution was evaporated to dryness on a rotary evaporator. The residue was added to ml. of water and extracted three times with 25 ml. each of methylene chloride. The extract was backwashed with 15 ml. of water, then dried over magnesium chloride and evaporated to dryness. The residue, 1.4 g., was distributed in a SOD-transfer counter current distribution using the solvent system, waterzethyl acetatezethanolzcyclohexane (1:1:1:1), and a single peak which fit the theoretical was observed at K=1.05. The material in tubes 240 to 280 was isolated as a syrup.

Analysis.Calcd. for C H N O S: C, 56.47; H, 8.58; N, 6.27; S, 7.18. Found: C, 56.24; H, 8.54; N, 6.13; S,

Thin layer chromatography (TLC) showed that this material consisted of two substances. One was 3,4-O-isopropylidenelincomycin; the other, 3,4-O-isopropylideneepilincomycin, moved slightly slower.

(D) EPILINCOMYCIN The syrup from Part C was stored at room temperature 5 hrs. in a solution containing 60 ml. of 0.25 N hydrochloric acid and ml. of ethanol. It was then kept at 0 C. for 4 days. Following neutralization with sodium bicarbonate, it was evaporated to 25 ml., then extracted with chloroform. The extract was washed with a little water and dried over magnesium sulfate, then evaporated to a residue. Thin layer chromatography of the residue showed two substances, both of which were active against S. lutea. The residue was chromatographed on a 14" X Florisil (a synthetic silicate of the type described in US. Patent 2,393,625) column which was eluted gradiently with solvent which varied continuously from 100% Skellysolve B (technical hexane) to 100% acetone. The total volume was 5000 ml. The two compounds were thus separated.

Fraction I: Tubes 5365 (40 ml. cuts) Epilincomycin. Assay 450 meg/ml.

Analysis.Calcd. for C H N O S: C, 50.92; H, 8.55; N, 6.60; S, 7.56. Found: C, 50.19; H, 7.91; N, 6.05; S, 6.42.

24 Fraction II. Tubes 73-104. Lincomycin. Assay 950 meg/mg.

(E) TCIILORO-Y-DEOXYEPILINCOMYCIN-HC1 Cl-i- XXXII 7-epilincomycin hydrochloride (0.85 g.) was suspended in 17 ml. of carbon tetrachloride. Thionyl chloride (4.5 ml.) was added and the reaction mixture stirred at 25 for 15 minutes. The solid dissolved to give a clear, colorless solution. The reaction was then heated at reflux for 2 hours, cooled to 25, evaporated under vacuum to a solid yellow residue. This crude product was dissolved in 10 ml. of ethanol, made basic with 0.1 N sodium hydroxide, diluted to 500 ml. with water and extracted four times with 50-m1. portions of chloroform. The combined chloroform extracts were back-washed two times with 20-ml. portions of saturated sodium chloride solution, filtered and evaporated under vacuum. The solid residue was slurried with 500 ml. of ether, filtered and hydrogen chloride gas passed into the filtrate. The solid precipitate was collected, dissolved in 3 m1. of ethanol, and re-precipitated by the addition of ether.

The solid was collected and dried. A 30% yield (270 mg.) of 7-chloro-7-deoxyepilincomycin assaying about 2 to 3 times the activity of lincomycin was obtained. TLC on silica gel (MeOI-I:CHCl system, 1:6 v./v.) showed one spot having an Rf of 0.44 as compared with an Rf of 0.52 for 7-chloro-7-deoxylincomycin.

Following the procedure of Preparation 1, Part A-2, 7-bromo-7-deoxylincomycin is obtained.

By substituting lincomycin in this preparation by lincomycin analogs of Formula II wherein Z, R, R R R of the Ac group are as given and illustrated above, the corresponding 7-halo-7-deoxyepi1incomycin analogs of the formula:

AcNli OH XXXIII wherein X is halogen and Z, R, R R and R of the Ac group are as given and illustrated above, are obtained. All the compounds that have been described above, therefore, have their counterpart in the opposite configuration, that is, the configuration derived from the 7-epi form. If an inversion is efiected by the substitution of the 7-hydroxy group by halogen, then the epi-compounds which have the L-threo configuration, are inverted to the Derythro configuration. In any event both the D-erythro and the L-threo forms are obtained, the one when the normal lincomycins (D-erythro) are used, the other when epi-lincomycins (L-threo) are used. Both forms, however, yield the same 6,7,8-trideoxy compound on halogenation.

While the processes of the invention have been described with reference to the preparation of specific compounds, it is to be understood that the process is broadly applicable to compounds of Formula II wherein Ac and R are radicals which are not reactive with thionyl chloride. Thus, in the broad process according to the invention, Ac can be hydrogen or any acyl that is non-reactive with thionyl chloride and R can be any alkyl, irrespec- AcNH- wherein R is alkyl of not more than 20 carbon atoms and Ac is the acyl radical of a 4-substituted-L-2- pyrrolidinecarboxylic acid of the formula selected from the group consisting of $112 Illa N N w m K a i OH Rx ll HR1 H O O wherein R and R are selected from the group consisting of alkylidene of not more than 20 carbon atoms, cycloalkylidene of from 3 to not more than 8 carbon atoms, and aralkylidene of not more than 12 carbon atoms; and R is selected from the group consisting of hydrogen and HR 2. A compound of the formula lliRn K NH HR1 H 26 wherein R and HR are lower alkyl and HR is hydrogen or lower alkyl.

3. A compound according to claim 2 wherein R is methyl or ethyl, HR is propyl, and HRg is hydrogen, methyl, or ethyl.

4. A compound according to claim 2 wherein R is methyl or ethyl, HR; is ethyl, and HRg is hydrogen, methyl, or ethyl.

5. A compound according to claim 2 wherein R is propyl or butyl, HR is propyl, and HR is hydrogen, methyl, or ethyl.

6. A compound according to claim 2 wherein R is propyl, HR is ethyl, and HR is hydrogen, meth or ethyl.

7. A compound according to claim 2 wherein R is methyl or ethyl, HR is pentyl, and HR is hydrogen, methyl, or ethyl.

8. A compound according to claim 2 wherein R is methyl or ethyl, HR is butyl, and HR is hydrogen, methyl, or ethyl.

9. A compound according to claim 2 wherein R is propyl or butyl, HR is butyl, and HR;,, is hydrogen, methyl, or ethyl.

10. A compound according to claim 2 wherein R is o cyclohexyl, HR is propyl, butyl, or pentyl, and HR; is

hydrogen, methyl, or ethyl. 11. 7-deoxylincomycin.

References Cited UNITED STATES PATENTS 2/1967 Magerlein 26O210 OTHER REFERENCES Pigman: The Carbohydrates, 1957, pp. 130, 373- 375, Academic Press Inc., New York, N.Y.

LEWIS GOTTS, Primary Examiner.

JOHNNIE R. BROWN, Assistant Examiner.

U.S. Cl. X.R. 

